Loss of Methylation at H19 DMD Is Associated with Biallelic Expression and Reduced Development in Cattle Derived by Somatic Cell Nuclear Transfer

Although cloning of mammals has been achieved successfully, the percentage of live offspring is very low because of reduced fetal size and fewer implantation sites. Recent studies have attributed such pathological conditions to abnormal reprogramming of the donor cell used for cloning. The inability of the oocyte to fully restore the differentiated status of a somatic cell to its pluripotent and undifferentiated state is normally evidenced by aberrant DNA methylation patterns established throughout the genome during development to blastocyst. These aberrant methylation patterns are associated with abnormal expression of imprinted genes, which among other genes are essential for normal embryo development and gestation. We hypothesized that embryo loss and low implantation rates in cattle derived by somatic cell nuclear transfer (SCNT) are caused by abnormal epigenetic reprogramming of imprinted genes. To verify our hypothesis, we analyzed the parental expression and the differentially methylated domain (DMD) methylation status of the H19 gene. Using a parental-specific analysis, we confirmed for the first time that H19 biallelic expression is tightly associated with a severe demethylation of the paternal H19 DMD in SCNT embryos, suggesting that these epigenetic anomalies to the H19 locus could be directly responsible for the reduced size and low implantation rates of cloned embryos in cattle.

Circadian Pattern of Wheel-Running Activity of a South American Subterranean Rodent (Ctenomys cf knightii)

Circadian rhythms are regarded as essentially ubiquitous features of animal behavior and are thought to confer important adaptive advantages. However, although circadian systems of rodents have been among the most extensively studied, most comparative biology is restricted to a few related species. In this study, the circadian organization of locomotor activity was studied in the subterranean, solitary north Argentinean rodent, Ctenomys knightii. The genus, Ctenomys, commonly known as Tuco-tucos, comprises more than 50 known species over a range that extends from 12S latitude into Patagonia, and includes at least one social species. The genus, therefore, is ideal for comparative and ecological studies of circadian rhythms. Ctenomys knightii is the first of these to be studied for its circadian behavior. All animals were wild caught but adapted quickly to laboratory conditions, with clear and precise activity-rest rhythms in a light-dark (LD) cycle and strongly nocturnal wheel running behavior. In constant dark (DD), the rhythm expression persisted with free-running periods always longer than 24h. Upon reinstatement of the LD cycle, rhythms resynchronized rapidly with large phase advances in 7/8 animals. In constant light (LL), six animals had free-running periods shorter than in DD, and 4/8 showed evidence of splitting. We conclude that under laboratory conditions, in wheel-running cages, this species shows a clear nocturnal rhythmic organization controlled by an endogenous circadian oscillator that is entrained to 24h LD cycles, predominantly by light-induced advances, and shows the same interindividual variable responses to constant light as reported in other non-subterranean species. These data are the first step toward understanding the chronobiology of the largest genus of subterranean rodents.

Morphology and phylogenetic relationships of a remarkable new genus and two new species of Neotropical freshwater stingrays from the Amazon basin (Chondrichthyes: Potamotrygonidae)

The morphology and phylogenetic relationships of a new genus and two new species of Neotropical freshwater stingrays, family Potamotrygonidae, are investigated and described in detail. The new genus, Heliotrygon, n. gen., and its two new species, Heliotrygon gomesi, n. sp. (type-species) and Heliotrygon rosai, n. sp., are compared to all genera and species of potamotrygonids, based on revisions in progress. Some of the derived features of Heliotrygon include its unique disc proportions (disc highly circular, convex anteriorly at snout region, its width and length very similar), extreme subdivision of suborbital canal (forming a complex honeycomb-like pattern anterolaterally on disc), stout and triangular pelvic girdle, extremely reduced caudal sting, basibranchial copula with very slender and acute anterior extension, and precerebral and frontoparietal fontanellae of about equal width, tapering very little posteriorly. Both new species can be distinguished by their unique color patterns: Heliotrygon gomesi is uniform gray to light tan or brownish dorsally, without distinct patterns, whereas Heliotrygon rosai is characterized by numerous white to creamy-white vermiculate markings over a light brown, tan or gray background color. Additional proportional characters that may further distinguish both species are also discussed. Morphological descriptions are provided for dermal denticles, ventral lateral-line canals, skeleton, and cranial, hyoid and mandibular muscles of Heliotrygon, which clearly corroborate it as the sister group of Paratrygon. Both genera share numerous derived features of the ventral lateral-line canals, neurocranium, scapulocoracoid, pectoral basals, clasper morphology, and specific patterns of the adductor mandibulae and spiracularis medialis muscles. Potamotrygon and Plesiotrygon are demonstrated to share derived characters of their ventral lateral-line canals, in addition to the presence of angular cartilages. Our morphological phylogeny is further corroborated by a molecular phylogenetic analysis of cytochrome b based on four sequences (637 base pairs in length), representing two distinct haplotypes for Heliotrygon gomesi. Parsimony analysis produced a single most parsimonious tree revealing Heliotrygon and Paratrygon as sister taxa (boot-strap proportion of 70%), which together are the sister group to a clade including Plesiotrygon and species of Potamotrygon. These unusual stingrays highlight that potamotrygonid diversity, both in terms of species composition and undetected morphological and molecular patterns, is still poorly known.

Potamotrygon tigrina, a new species of freshwater stingray from the upper Amazon basin, closely related to Potamotrygon schroederi Fernandez-Yepez, 1958 (Chondrichthyes: Potamotrygonidae)

A new species of Neotropical freshwater stingray, family Potamotrygonidae, is described from the Rio Nanay in the upper Rio Amazonas basin of Peru. Potamotrygon tigrina, n. sp., is easily distinguished from all congeners by its conspicuous dorsal disc coloration, composed of bright yellow to orange vermiculations strongly interwoven with a dark-brown to deep-black background. Additional features that in combination diagnose P. tigrina, n. sp., include the presence of a single angular cartilage, low and not closely grouped dorsal tail spines, and coloration of tail composed of relatively wide and alternating bands of creamy white and dark brown to black. Potamotrygon tigrina is closely related to Potamotrygon schroederi Fernandez-Yepez, 1958, which occurs in the Rio Negro (Brazil) and Rio Orinoco (Venezuela, Colombia). Both species are very similar in proportions and counts, and share features hypothesized to be derived within Potamotrygonidae, related to their specific angular cartilage morphology, distal tail color, dorsal tail-spine pattern, and ventral lateral-line system. To further substantiate the description of P. tigrina, n. sp., we provide a redescription of P. schroederi based on material from the Rio Negro (Brazil) and Rio Orinoco (Venezuela). Specimens from the two basins differ in number of vertebral centra and slightly in size and frequency of rosettes on dorsal disc, distinctions that presently do not warrant their specific separation. Potamotrygon tigrina is frequently commercialized in the international aquarium trade but virtually nothing is known of its biology or conservation status.

Prolonged exposure to palmitate impairs fatty acid oxidation despite activation of AMP-activated protein kinase in skeletal muscle cells

The aim of this study was to investigate the chronic effects of palmitate on fatty acid (FA) oxidation, AMPK/ACC phosphorylation/activation, intracellular lipid accumulation, and the molecular Mechanisms involved in these processes in skeletal muscle cells. Exposure of L6 myotubes for 8 h to 200, 400, 600, and 800 mu M of palmitate did rot affect cel viability but significantly reduced FA oxidation by similar to 26.5%, similar to 43.5%, similar to 50%, and similar to 47%, respectively. Interestingly, this occurred despite significant increases in AMPK (similar to 2.5-fold) and ACC (similar to 3-fold) phosphorylation and in malonyl-CoA decarboxylase activity (similar to 38-60%). Low concentrations of palmitate (50-100 mu M) caused an increase (similar to 30%) in CPT-I activity. However, as the concentration of palmitate increased, CPT-I activity decreased by similar to 32% after exposure for 8 h to 800 mu M of palmitate. Although FA uptake was reduced (similar to 35%) in cells exposed to increasing, palmitate concentrations, intracellular lipid accumulation increased in a dose-dependent manner, reaching values similar to 2.3-, similar to 3-, and 4-fold higher than control in muscle cells exposed to 400, 600, and 800 mu M palmitate, respectively. Interestingly, myotubes exposed to 400 mu M of palmitate for 1h increased basal glucose uptake and glycogen synthesis by similar to 40%. However, as time of incubation in the presence of palmitate progressed from 1 to 8h, these increases were abolished and a time-dependent inhibition of insulin-stimulated glucose uptake (similar to 65%) and glycogen synthesis (30%) was observed in myotubes. These findings may help explain the dysfunctional adaptations that occur in glucose and FA Metabolism in skeletal muscle under conditions of chronically elevated circulating levels of non-esterified FAs. Such as in obesity and Type 2 Diabetes.

Effects of Exercise Training on Hepatic Microsomal Triglyceride Transfer Protein Content in Rats

Microsomal triglyceride transfer protein (MTP) is a protein that exerts a central regulatory role in very-low-density lipoprotein (VLDL) assembly and secretion. The purpose of the study was to investigate the effects of all exercise-training program oil hepatic content of MTP and its relation to hepatic VLDL-triglyceride (VLDL-TG) production in response to lipid infusion. Female rats either fed a standard (SD) or all obesity-induced high-fat (HF; 43% as energy) diet for 8 weeks were Subdivided into sedentary (Sed) and trained (Tr) groups. Exercise training consisted Of Continuous running on a motor-driven rodent treadmill 5 times/week for 8 weeks. At the end of this period, all rats in the fasted state were intravenously infused with a 20% Solution of intralipid for 3 h followed by all injection of Triton WR1339 to block lipoprotein lipase. An additional control grout) consisting of Sed rats fed the SD diet was infused with saline (0.9% NaCl). Plasma TG accumulation was thereafter measured during 90 min to estimate VLDL-TG production. Under HF diet, hepatic MTP content and plasma TG accumulation after Triton blockade (thus reflecting VLDL-TG synthesis and secretion) were not changed in Sed rats, whereas liver TG content was highly increased (similar to 90%; p<0.01). Oil the other hand, training reduced liver MTP protein content in both SD(-18%) and HF(-23%) fed rats(p<0.05). Plasma VLDL-TG accumulation was also lower (p<0.05) in Tr than in Sed rats fed the HF diet. This effect was not observed in SD fed rats. Furthermore, the exercise training-induced decrease in VLDL-TG production in HF rats was associated with a decrease in liver TG levels. It is Concluded that in addition to a reduction in liver TG content, exercise training reduces VLDL synthesis and/or secretion in HF fed rats probably via MTP regulation.

Checking Completeness of Tests for Finite State Machines

In testing from a Finite State Machine (FSM), the generation of test suites which guarantee full fault detection, known as complete test suites, has been a long-standing research topic. In this paper, we present conditions that are sufficient for a test suite to be complete. We demonstrate that the existing conditions are special cases of the proposed ones. An algorithm that checks whether a given test suite is complete is given. The experimental results show that the algorithm can be used for relatively large FSMs and test suites.

Fault Coverage-Driven Incremental Test Generation

In this paper, we consider a classical problem of complete test generation for deterministic finite-state machines (FSMs) in a more general setting. The first generalization is that the number of states in implementation FSMs can even be smaller than that of the specification FSM. Previous work deals only with the case when the implementation FSMs are allowed to have the same number of states as the specification FSM. This generalization provides more options to the test designer: when traditional methods trigger a test explosion for large specification machines, tests with a lower, but yet guaranteed, fault coverage can still be generated. The second generalization is that tests can be generated starting with a user-defined test suite, by incrementally extending it until the desired fault coverage is achieved. Solving the generalized test derivation problem, we formulate sufficient conditions for test suite completeness weaker than the existing ones and use them to elaborate an algorithm that can be used both for extending user-defined test suites to achieve the desired fault coverage and for test generation. We present the experimental results that indicate that the proposed algorithm allows obtaining a trade-off between the length and fault coverage of test suites.

Regulation of Radial Glial Motility by Visual Experience

Radial glia in the developing optic tectum express the key guidance molecules responsible for topographic targeting of retinal axons. However, the extent to which the radial glia are themselves influenced by retinal inputs and visual experience remains unknown. Using multiphoton live imaging of radial glia in the optic tectum of intact Xenopus laevis tadpoles in conjunction with manipulations of neural activity and sensory stimuli, radial glia were observed to exhibit spontaneous calcium transients that were modulated by visual stimulation. Structurally, radial glia extended and retracted many filopodial processes within the tectal neuropil over minutes. These processes interacted with retinotectal synapses and their motility was modulated by nitric oxide (NO) signaling downstream of neuronal NMDA receptor (NMDAR) activation and visual stimulation. These findings provide the first in vivo demonstration that radial glia actively respond both structurally and functionally to neural activity, via NMDAR-dependent NO release during the period of retinal axon ingrowth.

Controlled fabrication of gold nanoparticles biomediated by glucose oxidase immobilized on chitosan layer-by-layer films

The control of size and shape of metallic nanoparticles is a fundamental goal in nanochemistry, and crucial for applications exploiting nanoscale properties of materials. We present here an approach to the synthesis of gold nanoparticles mediated by glucose oxidase (GOD) immobilized on solid substrates using the Layer-by-Layer (LbL) technique. The LbL films contained four alternated layers of chitosan and poly(styrene sulfonate) (PSS), with GOD in the uppermost bilayer adsorbed on a fifth chitosan layer: (chitosan/PSS)(4)/(chitosan/GOD). The films were inserted into a solution containing gold salt and glucose, at various pHs. Optimum conditions were achieved at pH 9, producing gold nanoparticles of ca. 30 nm according to transmission electron microscopy. A comparative study with the enzyme in solution demonstrated that the synthesis of gold nanoparticles is more efficient using immobilized GOD. (C) 2009 Elsevier B.V. All rights reserved.

Involutions of RA Loops

Let L be an RA loop, that is, a loop whose loop ring over any coefficient ring R is an alternative, but not associative, ring. Let l bar right arrow l(theta) denote an involution on L and extend it linearly to the loop ring RL. An element alpha is an element of RL is symmetric if alpha(theta) = alpha and skew-symmetric if alpha(theta) = -alpha. In this paper, we show that there exists an involution making the symmetric elements of RL commute if and only if the characteristic of R is 2 or theta is the canonical involution on L, and an involution making the skew-symmetric elements of RL commute if and only if the characteristic of R is 2 or 4.

POLYNOMIAL AND GROUP IDENTITIES IN ALTERNATIVE LOOP ALGEBRAS

We investigate polynomial identities on an alternative loop algebra and group identities on its (Moufang) unit loop. An alternative loop ring always satisfies a polynomial identity, whereas whether or not a unit loop satisfies a group identity depends on factors such as characteristic and centrality of certain kinds of idempotents.

On E-functions of semisimple Lie groups

We develop and describe continuous and discrete transforms of class functions on a compact semisimple, but not simple, Lie group G as their expansions into series of special functions that are invariant under the action of the even subgroup of the Weyl group of G. We distinguish two cases of even Weyl groups-one is the direct product of even Weyl groups of simple components of G and the second is the full even Weyl group of G. The problem is rather simple in two dimensions. It is much richer in dimensions greater than two-we describe in detail E-transforms of semisimple Lie groups of rank 3.

Some classes of semisimple group (and loop) algebras over finite fields

We determine the structure of the semisimple group algebra of certain groups over the rationals and over those finite fields where the Wedderburn decompositions have the least number of simple components We apply our work to obtain similar information about the loop algebras of mdecomposable RA loops and to produce negative answers to the isomorphism problem over various fields (C) 2010 Elsevier Inc All rights reserved

Paper Microzone Plates

This paper describes 96- and 384-microzone plates fabricated in paper as alternatives to conventional multi-well plates fabricated in molded polymers. Paper-based plates are functionally related to plastic well plates, but they offer new capabilities. For example, paper-microzone plates are thin (similar to 180 mu m), require small volumes of sample (5 mu L per zone), and can be manufactured from inexpensive materials ($0.05 per plate). The paper-based plates are fabricated by patterning sheets of paper, using photolithography, into hydrophilic zones surrounded by hydrophobic polymeric barriers. This photolithography used an inexpensive formulation photoresist that allows rapid (similar to 15 min) prototyping of paper-based plates. These plates are compatible with conventional microplate readers for quantitative absorbance and fluorescence measurements. The limit of detection per zone loaded for fluorescence was 125 fmol for fluorescein isothiocyanate-labeled bovine serum albumin, and this level corresponds to 0.02 the quantity of analyte per well used to achieve comparable signal-to-noise in a 96-well plastic plate (using a solution of 25 nM labeled protein). The limits of detection for absorbance on paper was aproximately 50 pmol per zone for both Coomassie Brilliant Blue and Amaranth dyes; these values were 0.4 that required for the plastic plate. Demonstration of quantitative colorimetric correlations using a scanner or camera to image the zones and to measure the intensity of color, makes it possible to conduct assays without a microplate reader.